On a worldwide basis, leptospirosis is the most widespread zoonotic disease. Leptospirosis is also emerging as a significant infectious disease in urban slums, particularly in tropical regions. The infection is caused by spirochetes of the genus Leptospira, and varies in severity from mild illness to fatal hemorrhagic disease with multiple organ failure. Disease severity varies with the host species, host age and health status, and the infecting strain. Virtually all species of mammals can serve as carriers, with the Leptospira persisting in the proximal tubules of the kidney and being shed in the urine. The persistence of leptospirosis in wildlife, companion animals, livestock, and in particular peridomestic rats, results in a continuous reservoir for human infection through exposure to animal tissue or body fluids containing Leptospira, or to urine-contaminated water sources. The bacteria enter through mucous membranes or skin abrasions, then disseminate to multiple tissues and can establish persistent infection in particular sites, such as the proximal tubules of the kidney. Adhesion to host cell surfaces and extracellular matrix (ECM) is critical to the pathogenesis of disease for many pathogens. This is likely to be true of Leptospira, as well, but most previous investigations into adhesion of Leptospira to mammalian tissue components have focused on interactions with the ECM. This proposal will focus on recent findings that the bacteria bind more efficiently to the cell surface than to the ECM, suggesting the presence of additional, significant, unexplored aspects of the interactions of pathogenic leptospires with the host. A portion of this adhesion activity is attributable to proteoglycans, which consist of glycosaminoglycans linked to core proteins. Chondroitin sulfate B proteoglycans are the primary receptors for L. interrogans in this class. Additional, non-proteoglycan receptors await identification. In addition, recent data show that L. interrogans, but not the saprophytic species L. biflexa, damages endothelial cell monolayers, as revealed by microarray analyses of transcript levels, staining of the actin cytoskeleton, and rounding of the cells. These changes lead to increased permeability of the endothelial layers to the pathogen, without evidence of necrosis or apoptosis. Therefore the goals of this project are to 1) Analyze the importance of proteoglycans in Leptospira interrogans attachment to proximal tubule epithelial cells, 2) Identify the non-proteoglycan mammalian cell surface receptor(s) for L. interrogans, 3) Identify Leptospira interrogans molecules involved in attachment to human cells, and 4) Explore the mechanism of damage to endothelial cells by Leptospira interrogans. The long-term goal of this project is to better understand the nature and consequences of Leptospira interactions with mammalian cells, which may eventually be useful in future development of novel vaccines or therapeutic approaches.